Neuropathic pain is common and disabling. Although understanding of pain mechanisms has advanced, available treatments remain inadequate. The overall goal of the proposed research is to devise a highly effective and flexible treatment system by combining two novel approaches. First, therapy that targets a single (or several) dorsal root ganglia (DRGs) will permit direct treatment of sensory neurons on a segmental basis, thereby limiting side effects on healthy sensory pathways. Second, transplantation of mesenchymal stem cells (MSCs) provides a powerful opportunity to capitalize on expanding pathophysiological understanding. Specifically, autologous MSCs are readily available, they are easily grown, they are highly capable of secreting peptides after genetic engineering, and there are no ethical or regulatory issues with these cells. Preliminary data indicate that in vitro lentiviral transduction produces stable transgene expression in MSCs through many cycles of propagation, since this vector inserts the transgene into the host genome, and that MSCs secrete much higher levels of therapeutic peptides than they can without genetic modification. Unlike gene therapy approaches using direct viral vector injection into patients, this strategy avoids immunogenic and toxic effects of the virus, and permits controlled, highly efficient transduction outside the patient prior to reimplantation, thereby enhancing performance and safety. The proposed work will develop this new therapy in sequential Aims. First, the necessary dose of MSCs, their survival rates, and their differentiation fate will be characterized using rats as subjects. Second, important anatomical and physiological observations will evaluate effects of MSC transplantation on endogenous DRG cells to confirm that implantation of MSCs per se has no harmful effects. Finally, proof of concept experiments will measure the efficacy of treating neuropathic pain in a rat nerve-injury model, by transplantation of engineered MSCs into the DRG. The modified MSCs used in these translational trials will secrete either glial cell derived neurotrophic factor (GDNF) or interleukin 10, which have demonstrated efficacy in preventing or reversing neuropathic pain when delivered by other routes. Pain behavior and cellular effects will be compared to animals in which DRGs have been injected with MSCs that express only a reporter gene (GFP), and to untreated controls. Completion of the proposed work will establish the basis for therapeutic trials of engineered MSC as sources of these or other analgesic peptides in larger animals or human subjects.